Nanoparticles are submicron materials that often possess different properties than bulk material of the same kind. Nanoparticles have been studied for uses in many fields, including diagnostic and therapeutic applications in the life sciences. Because of their small size and unique properties, nanoparticles often have enhanced distribution in the body compared to larger sized particles. Further, nanoparticles may be specifically directed to particular targets in the body by attaching one or more components to the nanoparticle surface (i.e. functionalization). Functionalization of a nanoparticle with a component having affinity for a specific target in the body, for example, collagen, can direct the nanoparticle to tissues containing the target molecule.
There are more than 20 types of collagen currently identified, with type I being the most common. Many tissues are composed primarily of type I collagen including tendon, ligament, skin, and bone. While each of these structures also contains other collagen types, proteoglycans and glycosaminoglycans, and minerals in the case of bone, the principle component is type I collagen.
Disclosed herein is targeted delivery of nanoparticles, utilizing collagen as an attachment site for functionalized nanoparticles. Collagens are a component of the extracellular matrix (ECM), which is responsible for supporting cells, thus making up connective tissues such as blood vessels, cartilage, or skin. Collagens and their corresponding binding domains have been well studied. Collagen binding domains can be reduced to specific peptide sequences, which bind specifically and exclusively to collagen, the target protein. Utilizing peptide synthesis, it is possible to immobilize a collagen-binding peptide, for example a collagen type I or type II binding peptide, onto nanoparticles, allowing those nanoparticles to fasten specifically to type I or type II collagen, respectively.
This targeted delivery system is not limited to type I collagen, as many other collagen types have also been defined. As such, specific targeting of a variety connective tissues and proteins is also contemplated. Collagen-binding gold nanoparticles, for example, are useful for a variety of biomedical imaging techniques including SEM, TEM, confocal microscopy, MRI, and photoacoustic imaging. Further, radiopaque nanoparticles that are coupled with collagen binding proteins can be used in radiologic imaging such as CT scan or X-ray.
The following various embodiments are contemplated:
1) A composition comprising at least one collagen binding polypeptide coupled to a nanoparticle, wherein the polypeptide contains from 7 amino acids to 40 amino acids and wherein the polypeptide does not form a triple helix.
2) The composition of clause 1 wherein the nanoparticle further comprises a stabilizer.
3) The composition of clause 2 wherein the stabilizer is selected from the group consisting of a polyethylene glycol (PEG), a dextran, a peptide, an alkane-thiol, and an oligonucleotide-thiol.
4) The composition of clause 3 wherein the stabilizer is PEG.
5) The composition of any of clauses 1 to 4 wherein the polypeptide contains from 9 amino acids to 40 amino acids.
6) The composition of any of clauses 1 to 4 wherein the polypeptide contains from 9 amino acids to 30 amino acids.
7) The composition of any of clauses 1 to 4 wherein the polypeptide contains from 9 amino acids to 18 amino acids.
8) The composition of any of clauses 1 to 4 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), RLDGNEIKR (SEQ ID NO: 8), KELNVYT (SEQ ID NO: 9), KLWVLPK (SEQ ID NO: 10), CQDSETRTFY (SEQ ID NO: 11), AHEEISTTNEGVM (SEQ ID NO: 12), GLRSKSKKFRRPDIQYPDATDEDITSHM (SEQ ID NO: 13), GSITTIDVPWNV (SEQ ID NO: 14), and NGVFKYRPRYFLYKHAYFYPPLKRFPVQ (SEQ ID NO: 15).
9) The composition of any of clauses 1 to 4 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), and RLDGNEIKR (SEQ ID NO: 8).
10) The composition of any of clauses 1 to 4 wherein the polypeptide comprises the amino acid sequence GELYKXILY, wherein X is serine, cysteine, or threonine (SEQ ID NO: 16).
11) The composition of any of clauses 1 to 4 wherein the polypeptide is RRANAALKAGELYKSILY (SEQ ID NO: 1).
12) The composition of any of clauses 1 to 4 wherein the polypeptide is RRANAALKAGELYKCILY (SEQ ID NO: 2).
13) The composition of any of clauses 1 to 12 further comprising a cysteine at the amino terminal region or carboxy terminal region.
14) The composition of clause 13 further comprising a spacer.
15) The composition of clause 14 wherein the spacer comprises at least one glycine.
16) The composition of any of clauses 1 to 12 further comprising a peptide sequence selected from the group consisting of GC, CG, and GCG.
17) The composition of any of clauses 1 to 16 wherein the nanoparticle is a polymeric nanoparticle, a metallic nanoparticle, a semiconductor nanoparticle, or any combination thereof.
18) The composition of clause 17 wherein the nanoparticle is a gold nanoparticle.
19) The composition of clause 17 wherein the nanoparticle is an iron oxide nanoparticle.
20) The composition of any of clauses 1 to 19 further comprising a carrier.
21) The composition of clause 20 wherein the carrier is a pharmaceutically acceptable carrier.
22) The composition of clause 21 wherein the carrier is a liquid carrier and is selected from the group consisting of saline, glucose, alcohols, glycols, esters, amides, and a combination thereof.
23) An effective dose of the composition of any of clauses 1 to 22 for administration to a patient, wherein the effective dose ranges from about 1 ng to about 1 mg per kilogram of body weight.
24) An effective dose of the composition of any of clauses 1 to 22 for administration to a patient, wherein the effective dose ranges from about 1 pg to about 10 ng per kilogram of body weight.
25) An effective dose of the composition of any of clauses 1 to 22 for administration to a patient, wherein the effective dose ranges from about 1 μg to about 100 μg per kilogram of body weight.
26) A composition for use in imaging a collagenous matrix, the composition comprising at least one collagen binding polypeptide coupled to a nanoparticle, wherein the polypeptide contains from 7 amino acids to 40 amino acids and wherein the polypeptide does not form a triple helix.
27) The composition of clause 26 wherein the nanoparticle further comprises a stabilizer.
28) The composition of clause 27 wherein the stabilizer is selected from the group consisting of a polyethylene glycol (PEG), a dextran, a peptide, an alkane-thiol, and an oligonucleotide-thiol.
29) The composition of clause 28 wherein the stabilizer is PEG.
30) The composition of any of clauses 26 to 29 wherein the polypeptide contains from 9 amino acids to 40 amino acids.
31) The composition of any of clauses 26 to 29 wherein the polypeptide contains from 9 amino acids to 30 amino acids.
32) The composition of any of clauses 26 to 29 wherein the polypeptide contains from 9 amino acids to 18 amino acids.
33) The composition of any of clauses 26 to 29 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), RLDGNEIKR (SEQ ID NO: 8), KELNVYT (SEQ ID NO: 9), KLWVLPK (SEQ ID NO: 10), CQDSETRTFY (SEQ ID NO: 11), AHEEISTTNEGVM (SEQ ID NO: 12), GLRSKSKKFRRPDIQYPDATDEDITSHM (SEQ ID NO: 13), GSITTIDVPWNV (SEQ ID NO: 14), and NGVFKYRPRYFLYKHAYFYPPLKRFPVQ (SEQ ID NO: 15).
34) The composition of any of clauses 26 to 29 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), and RLDGNEIKR (SEQ ID NO: 8).
35) The composition of any of clauses 26 to 29 wherein the polypeptide comprises the amino acid sequence GELYKXILY, wherein X is serine, cysteine, or threonine (SEQ ID NO: 16).
36) The composition of any of clauses 26 to 29 wherein the polypeptide is RRANAALKAGELYKSILY (SEQ ID NO: 1).
37) The composition of any of clauses 26 to 29 wherein the polypeptide is RRANAALKAGELYKCILY (SEQ ID NO: 2).
38) The composition of any of clauses 26 to 37 further comprising a cysteine at the amino terminal region or carboxy terminal region.
39) The composition of clause 38 further comprising a spacer.
40) The composition of clause 39 wherein the spacer comprises at least one glycine.
41) The composition of any of clauses 26 to 37 further comprising a peptide sequence selected from the group consisting of GC, CG, and GCG
42) The composition of any of clauses 26 to 41 wherein the nanoparticle is a polymeric nanoparticle, a metallic nanoparticle, a semiconductor nanoparticle, or any combination thereof.
43) The composition of clause 42 wherein the nanoparticle is a gold nanoparticle.
44) The composition of clause 42 wherein the nanoparticle is an iron oxide nanoparticle.
45) The composition of any of clauses 26 to 44 further comprising a carrier.
46) The composition of clause 45 wherein the carrier is a pharmaceutically acceptable carrier.
47) The composition of clause 46 wherein the carrier is a liquid carrier and is selected from the group consisting of saline, glucose, alcohols, glycols, esters, amides, and a combination thereof.
48) An effective dose of the composition of any of clauses 26 to 47 for administration to a patient, wherein the effective dose ranges from about 1 ng to about 1 mg per kilogram of body weight.
49) An effective dose of the composition of any of clauses 26 to 47 for administration to a patient, wherein the effective dose ranges from about 1 pg to about 10 ng per kilogram of body weight.
50) An effective dose of the composition of any of clauses 26 to 47 for administration to a patient, wherein the effective dose ranges from about 1 μg to about 100 μg per kilogram of body weight.
51) A method for imaging a collagenous matrix, the method comprising the steps of contacting a collagenous matrix with a composition comprising at least one collagen binding polypeptide coupled to a nanoparticle, wherein the polypeptide contains from 7 amino acids to 40 amino acids and wherein the polypeptide does not form a triple helix, and imaging the collagenous matrix.
52) The method of clause 51 wherein the nanoparticle further comprises a stabilizer.
53) The method of clause 52 wherein the stabilizer is selected from the group consisting of a polyethylene glycol (PEG), a dextran, a peptide, an alkane-thiol, and an oligonucleotide-thiol.
54) The method of clause 53 wherein the stabilizer is PEG.
55) The method of any of clauses 51 to 54 wherein the polypeptide contains from 9 amino acids to 40 amino acids.
56) The method of any of clauses 51 to 54 wherein the polypeptide contains from 9 amino acids to 30 amino acids.
57) The method of any of clauses 51 to 54 wherein the polypeptide contains from 9 amino acids to 18 amino acids.
58) The method of any of clauses 51 to 54 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), RLDGNEIKR (SEQ ID NO: 8), KELNVYT (SEQ ID NO: 9), KLWVLPK (SEQ ID NO: 10), CQDSETRTFY (SEQ ID NO: 11), AHEEISTTNEGVM (SEQ ID NO: 12), GLRSKSKKFRRPDIQYPDATDEDITSHM (SEQ ID NO: 13), GSITTIDVPWNV (SEQ ID NO: 14), and NGVFKYRPRYFLYKHAYFYPPLKRFPVQ (SEQ ID NO: 15).
59) The method of any of clauses 51 to 54 wherein the polypeptide is selected from the group consisting of RRANAALKAGELYKSILY (SEQ ID NO: 1), RRANAALKAGELYKCILY (SEQ ID NO: 2), GELYKSILY (SEQ ID NO: 3), GELYKCILY (SEQ ID NO: 4), SYIRIADTNIT (SEQ ID NO: 5), TKKTLRT (SEQ ID NO: 6), SQNPVQP (SEQ ID NO: 7), and RLDGNEIKR (SEQ ID NO: 8).
60) The method of any of clauses 51 to 54 wherein the polypeptide comprises the amino acid sequence GELYKXILY, wherein X is serine, cysteine, or threonine (SEQ ID NO: 16).
61) The method of any of clauses 51 to 54 wherein the polypeptide is RRANAALKAGELYKSILY (SEQ ID NO: 1).
62) The method of any of clauses 51 to 54 wherein the polypeptide is RRANAALKAGELYKCILY (SEQ ID NO: 2).
63) The composition of any of clauses 51 to 62 further comprising a cysteine at the amino terminal region or carboxy terminal region.
64) The composition of clause 63 further comprising a spacer.
65) The composition of clause 64 wherein the spacer comprises at least one glycine.
66) The composition of any of clauses 51 to 62 further comprising a peptide sequence selected from the group consisting of GC, CG, and GCG.
67) The method of any of clauses 51 to 66 wherein the nanoparticle is a polymeric nanoparticle, a metallic nanoparticle, a semiconductor nanoparticle, or any combination thereof.
68) The method of clause 67 wherein the nanoparticle is a gold nanoparticle.
69) The method of clause 67 wherein the nanoparticle is an iron oxide nanoparticle.
70) The method of any of clauses 51 to 69 further comprising a carrier.
71) The method of clause 70 wherein the carrier is a pharmaceutically acceptable carrier.
72) The method of clause 71 wherein the carrier is a liquid carrier and the liquid carrier is selected from the group consisting of saline, glucose, alcohols, glycols, esters, amides, and a combination thereof.
73) The method of any of clauses 51 to 72 wherein an effective dose is administered to a patient, the effective dose ranging from about 1 ng to about 1 mg per kilogram of body weight.
74) The method of any of clauses 51 to 72 wherein an effective dose is administered to a patient, the effective dose ranging from about 1 pg to about 10 ng per kilogram of body weight.
75) The method of any of clauses 51 to 72 wherein an effective dose is administered to a patient, the effective dose ranging from about 1 μg to about 100 μg per kilogram of body weight.
76) The composition or method of any of clauses 1 to 75 wherein the polypeptide is coupled to the nanoparticle using a crosslinking agent.